ABSTRACT: Sepsis is a ?life-threatening organ dysfunction caused by a dysregulated host response to infection?. Neutrophils become hyperactive during sepsis and they mediate much of the morbidity and mortality associated with the disease. However the antimicrobial function of neutrophils provides benefit such that depletion of the entire population of neutrophils in the septic patient would be immunosuppressive. Heterogeneity of neutrophils has recently been described in disease states including cancer and lupus. Our laboratories have discovered the presence of a small population of bloodstream neutrophils that overexpress the integrin VLA3 (CD49c/CD29; a3b1) in ICU patients and experimental animals with sepsis. We have shown that blockade or genetic depletion of neutrophil VLA3 has a survival advantage in septic mice. These findings indicate that selective removal of the VLA3high subset improves outcome without impairment of innate host defense against infection. In humans, neutrophils from septic patients demonstrate pronounced damage to the barrier function of endothelial cells in vitro which is prevented by VLA3 blocking. This suggests that the primary target of damage to the septic host by VLA3high cells is the vasculature. In addition, preliminary evidence also shows that septic patients that have VLA3high neutrophils late into the disease have increased mortality as compared to patients who do not show VLA3high cells in their blood. The overarching hypothesis to be tested in this proposal is that a distinct, hyperinflammatory and damaging subset of blood neutrophils, which express high levels of VLA3, arises in the circulation of ICU patients with sepsis. It is hypothesized further that the presence of circulating VLA3high neutrophils predicts severity of the disease as determined by clinical scoring, and that VLA3 represents a druggable target for attenuation of inflammatory damage to the septic patient while avoiding overt immunosuppression. Three specific aims are put forth to test these hypotheses. Aim 1 will determine the correlation between VLA3 expression and activation on peripheral blood neutrophils and illness severity in septic ICU patients. Aim 2 will use human cells and multiphoton imaging of septic mice to test the hypothesis that VLA3high neutrophils comprise a hyperinflammatory subset that causes vascular damage and capillary leak. Aim 3 will characterize the VLA3high and VLA3low neutrophil subsets and examine the mechanisms through which they impair endothelial barrier function. Taken together, this proposal addresses mechanisms regarding how the dysregulated host response threatens patient survival and offers a molecular target for dampening the destructive arms of the hyperinflammatory response while preserving the beneficial aspects of immune defense.